The present invention relates to IL-16 antagonists and the use thereof for the treatment of IL-16 mediated disorders such as certain inflammatory diseases. In particular, the present invention relates to the discovery of IL-16 antagonist peptides whose sequences coincide with the C-terminal region of IL-16.
Interleukin-16 (IL-16), previously named lymphocyte chemoattractant factor (or LCF), is a pro-inflammatory lymphokine with chemoattractant activity for resting CD4+ T lymphocytes. Subsequent studies indicate that IL-16 activates signal transduction in CD4+ target cells including monocytes, eosinophils and pro-B cells, and stimulates a variety of biological activities in addition to chemotaxis. Among these activities are inhibition of retroviral replication (Maciaszek, et al., J. Immunol. 158:5, 1997; Zhou, et al., Nature Medicine 3:659, 1997 and Baier, et al., Nature 378:563, 1995), upregulation of IL-2R and synergy with IL-2 for CD4+ T cell proliferation (Parada, et al., J. Immunol. 160:2115, 1998), induction of RAG-1 and RAG-2 expression in CD4+ pro-B cells (Szabo, et al., J. Immunol., 161:2248, 1998), and transient inhibition of Mixed Lymphocyte Reaction (MLR)(Theodore, et al., J. Immunol. 157:1958, 1996). Investigation of certain human diseases and experimental murine models indicates that IL-16 participates in inflammatory conditions characterized by tissue recruitment of CD4+T lymphocytes and other CD4+ cell types. Conditions where IL-16 has been identified by ELISA and/or bioassay of body fluids, or by immunohistochemical and in situ hybridization techniques, include bronchial asthma (Laherge et al., Am. J. Respir. Cell. Mol. Biol. 17: 193, 1997), inflammatory bowel disease (Keates et al., Gastroenterology 112, A110, 1997), Graves"" disease (Cruikshank et al., J. Allergy Clin. Immunol. 99: 554, 1997), multiple sclerosis (Biddison et al., J. Immunol. 158: 3046, 1997) and bullous pemphigoid (Center et al., J. Invest. Dermatol. 81: 204, 1983). IL-16 is also implicated in the pathogenesis of rheumatoid arthritis (Klimiuk et al., J. Immunol. 162: 4293-4299, 1999) and lupus (Lee et al., British J. Rheumatology 37: 1334-1337, 1998). Thus, it would be desirable to identify and/or generate reagents capable of interfering with the IL-16 activity for the purpose of treating inflammatory diseases.
The predicted amino acid sequence of IL-16 contains a central PDZ module, and structural studies confirm that IL-16 assumes a core PDZ-like conformation with flexible N-terminal and C-terminal tails of 17 and 14 residues, respectively (Muhlhahn et al., Nature Structural Biology 5:682, 1998)). A synthetic oligopeptide corresponding to the 16 C-terminal amino acids of human IL-16 (Arg106 to Ser121) has been reported to inhibit the chemoattractant activity of natural and recombinant human or murine IL-16 (Keane et al., J. Immunol. 160:5945, 1998).
The present invention demonstrates that a series of peptides corresponding to native or substituted sequences of the C-terminus of IL-16 can inhibit IL-16 activity. Compositions and methods useful for treating IL-16 mediated disorders are exploited using these peptides.
One embodiment of the present invention is directed to IL-16 antagonists.
Another embodiment of the present invention is directed to IL-16 antagonist peptides.
The IL-16 antagonist peptides of the present invention are at least 4 amino acids in length and substantially correspond to the C-terminal sequence of human or murine IL-16 surrounding the Arg/Lys-Arg motif, i.e., R106-R107 of human IL-16, R103-R104 of murine IL-16 or K106-R107 of IL-16 from squirrel monkey, for example.
A preferred IL-16 antagonist peptide of the present invention is the tetrameric peptide Xaa0RXaa1Xaa2 (SEQ ID NO:1), wherein Xaa0 is Arg or Lys, and Xaa1 and Xaa2 can be any amino acid. Preferably, Xaa1 and Xaa2 are those amino acids found in the native sequence of a mammalian IL-16, such as Lys or Thr for Xaa1, and Ser for Xaa2.
More preferably, Xaa0RXaa1Xaa2 is a tetramer having a sequence which coincides with the native sequence of a mammalian IL-16, e.g., RRKS (SEQ ID NO:2), RRTS (SEQ ID NO:3), or KRKS (SEQ ID NO:4). Even more preferably, such tetramer has Arg as the first amino acid. Homologs and analogs of the tetramers of SEQ ID NO:2-4 are also contemplated by the present invention. For example, analogs of RRKS (SEQ ID NO:2) include RRAS (SEQ ID NO:5) and RRKA (SEQ ID NO:6).
Another preferred IL-16 antagonist peptide of the present invention is a tetrameric peptide having the sequence of Xaa1Xaa2Xaa0R (SEQ ID NO:8), wherein Xaa0 is Arg or Lys, and Xaa1 and Xaa2 can be any amino acid.
Preferably, Xaa1 and Xaa2 are those amino acids found in the native sequence of a mammalian IL-16, e.g., Val for Xaa1, and Ile or Leu for Xaa2.
More preferably, Xaa1Xaa2Xaa0R is a tetramer having a sequence which coincides with the native sequence of a mammalian IL-16, such as VIRR (SEQ ID NO:9), VLRR (SEQ ID NO:10) and VIKR (SEQ ID NO:11). Even more preferably, such tetramer has Arg as the first amino acid. Homologs and analogs of these tetramers (SEQ ID NOS:9-11) are also contemplated by the present invention.
Still another preferred IL-16 antagonist peptide of the present invention is a tetrameric peptide having the sequence of Xaa1Xaa0RXaa2 (SEQ ID NO:12), wherein Xaa0 is Arg or Lys, and Xaa1 and Xaa2 can be any one amino acid. Preferably, Xaa1 and Xaa2 are those amino acids found in the native sequence of a mammalian IL-16. For example, Xaa1 can be Ile or Leu, and Xaa2 can be Lys or Thr.
More preferably, Xaa1Xaa2Xaa0R is a tetramer having a sequence which coincides with the native sequence of an IL-16, such as IRRK (SEQ ID NO:13), IRRT (SEQ ID NO:14), LRRK (SEQ ID NO:15), and IKRK (SEQ ID NO:16). Even more preferably, such tetramer has Arg as the first amino acid. Homologs and analogs of such tetramers are also contemplated by the present invention.
Further according to the present invention, an IL-16 antagonist peptide can be longer than a tetramer, as long as the such antagonist peptide contains one of the tetrameric sequences described hereinabove, i.e., Xaa0RXaa1Xaa2 (SEQ ID NO:1), Xaa1Xaa0RXaa2 (SEQ ID NO:8) or Xaa1Xaa2Xaa0R (SEQ ID NO:12), and as long as such peptide antagonizes at least one IL-16 biological activity.
Nucleic acid molecules coding for any of the above IL-16 antagonist peptide of the present invention, expression vectors which include any of such nucleic acid molecules, as well as related host cells containing such nucleotide sequences or vectors, are also contemplated by the present invention.
In a further aspect, the present invention provides antibodies directed against the IL-16 antagonist peptides of the present invention.
Preferably, the antibodies of the present invention are raised against those IL-16 antagonist peptides whose sequences coincide with the corresponding sequences of a mammalian IL-16 protein, which antibodies can antagonize or neutralize the activity of IL-16. Both polyclonal antibodies and monoclonal antibodies are contemplated by the present invention.
Functional derivatives of the monoclonal antibodies of the present invention are also contemplated, including Fab, Fabxe2x80x2, F(abxe2x80x2)2 of the present mAbs, single chain antibodies, humanized antibodies and the like.
A related aspect of the present invention is directed to methods of raising antibodies specific for the IL-16 antagonist peptides of the present invention by using such peptides as immunogens.
In another embodiment, the present invention provides pharmaceutical compositions which include one or more of the IL-16 antagonist peptides or antibodies, and a pharmaceutically acceptable carrier. The pharmaceutical compositions of the present invention can also include other appropriate active ingredients, such as known anti-inflammatory agents, e.g., anti-CD4 antibodies, anti-TNFxcex1 antibodies, NSAIDS, steroids, cyclosporin-A, or cytotoxic drugs.
Another aspect of the present invention provides methods of interfering with, blocking or otherwise preventing the interaction or binding of IL-16 with an IL-16 receptor by employing the IL-16 antagonists contemplated by the present invention.
In a further aspect, the present invention provides methods of treating an IL-16-mediated disorder in a subject by administering a therapeutically effective amount of a pharmaceutical composition of the present invention. In particular, IL-16-mediated disorders which can be treated by employing the methods of the present invention include asthma, rheumatoid arthritis, inflammatory bowel disease, Graves"" disease, multiple sclerosis, lupus and bullous pemphigoid.